GeneRead Pure mRNA Kit

For highly selective and efficient purification of mRNA for NGS applications

Efficient removal of almost all rRNAs (significantly >99%)

No bias from unintended removal of poly A+ mRNAs

As little as 100 ng – 5 μg total RNA required

Maximal convenience, minimal hands-on time

Automation on the QIAcube

The GeneRead Pure mRNA Kit allows highly specific polyA-oligo-dT–based purification of mRNA ready for next-generation sequencing (NGS) applications. By effectively depleting rRNA and non-adenylated, non-coding, as well as regulatory RNA from a wide variety of eukaryotic species, the kit enriches for mRNA, ensuring optimal use of expensive sequencing capacity and resources. With a preparation time of <45 min and convenient automation options, the GeneRead Pure mRNA Kit offers unparalleled convenience by minimizing the hands-on time required to purify mRNA. Procedure can be automated on the QIAcube.

The performance of the GeneRead Pure mRNA Kit and mRNA purification kits from other suppliers was compared. Starting material for mRNA purification was 0.5 μg total RNA. Despite only requiring one purification round, the GeneRead Pure mRNA Kit provides superior enrichment of mRNA. Residual rRNA is still visible in the form of two distinct bands in samples prepared using products from other suppliers.

Specific and efficient mRNA enrichment.

For analysis, 5 μg total RNA from Jurkat cells was treated using the GeneRead Pure mRNA protocol. Subsequently, libraries of the enriched samples, as well as of the total RNA were made using the same method and sequenced in multiplex by Illumina sequencing on a MiSeq system. Reads were mapped against the Biomart database and the percentage of mapped reads from each library for the biotypes was plotted. rRNAs are often listed under low qual. unmapped, as the annotation status of rRNA transcripts is often still suboptimal.

Convenient, streamlined procedure.

Performance

Exceptional convenience and cost-effectiveness

The GeneRead Pure mRNA Kit contains spin columns and all necessary reagents and buffers for the isolation of pure poly A+ mRNA from total RNA preparations in <45 minutes. GeneRead Pure mRNA technology combines the convenient handling of magnetic particles with the specificity of oligo-dT hybridization. The streamlined protocol only requires short pretreatment steps. Products from other suppliers involve more time-consuming procedures as they generally require a second round of purification to achieve mRNA pure enough for NGS applications, thereby resulting in increased resources and costs. The GeneRead Pure mRNA Kit is the only kit on the market that delivers mRNA of maximal purity with only one round of purification. Poly A+ mRNA purified with GeneRead Pure mRNA technology does not require further purification and is ready for downstream use in applications such as NGS. After purification with the GeneRead Pure mRNA Kit, only meaningful reads are left — the majority of these reads are from polyA+ mRNA.

Using the GeneRead Pure mRNA Kit, 5 μg total RNA from Jurkat cells was purified. To determine the efficacy of removal of different types of rRNA, quantitative RT-PCR was performed.

Principle

GeneRead Pure mRNA technology combines the convenient handling of magnetic particles with the specificity of oligo-dT hybridization. Protocols require short pretreatment steps. Oligo-dT probes are covalently attached to the surface of Pure mRNA Beads. mRNA binds rapidly and efficiently to the oligo-dT probes on the Pure mRNA Beads in the presence of Buffer mRBB (see figure Convenient, streamlined procedure). mRNA bound to the magnetic particles is then efficiently washed. Two wash steps are used to considerably improve the purity of the mRNA. Highly pure mRNA is eluted in the elution buffer (Buffer OEB) provided with the kit. mRNA yields depend on sample input and sample storage.

Minimal hands-on time required

The majority of the GeneRead Pure mRNA procedure — from hybridization and capture through to RNA cleanup — can be fully automated on the QIAcube, affording maximal convenience and greater processing flexibility, and minimizing the hands-on time required.

Applications

RNA prepared using the GeneRead Pure mRNA Kit is suited for use in a wide range of downstream applications, such as: